scholarly journals Targeting the coronavirus nucleocapsid protein through GSK-3 inhibition

2021 ◽  
Vol 118 (42) ◽  
pp. e2113401118
Author(s):  
Xiaolei Liu ◽  
Anurag Verma ◽  
Gustavo Garcia ◽  
Holly Ramage ◽  
Anastasia Lucas ◽  
...  
Keyword(s):  
Nucleocapsid Protein ◽  
Glycogen Synthase ◽  
Lung Epithelial Cells ◽  
Dependent Manner ◽  
Virion Assembly ◽  
N Protein ◽  
Consensus Sequences ◽  
Lung Epithelial ◽  
A Cell ◽  
Transcription And Replication

The coronaviruses responsible for severe acute respiratory syndrome (SARS-CoV), COVID-19 (SARS-CoV-2), Middle East respiratory syndrome-CoV, and other coronavirus infections express a nucleocapsid protein (N) that is essential for viral replication, transcription, and virion assembly. Phosphorylation of N from SARS-CoV by glycogen synthase kinase 3 (GSK-3) is required for its function and inhibition of GSK-3 with lithium impairs N phosphorylation, viral transcription, and replication. Here we report that the SARS-CoV-2 N protein contains GSK-3 consensus sequences and that this motif is conserved in diverse coronaviruses, raising the possibility that SARS-CoV-2 may be sensitive to GSK-3 inhibitors, including lithium. We conducted a retrospective analysis of lithium use in patients from three major health systems who were PCR-tested for SARS-CoV-2. We found that patients taking lithium have a significantly reduced risk of COVID-19 (odds ratio = 0.51 [0.35–0.74], P = 0.005). We also show that the SARS-CoV-2 N protein is phosphorylated by GSK-3. Knockout of GSK3A and GSK3B demonstrates that GSK-3 is essential for N phosphorylation. Alternative GSK-3 inhibitors block N phosphorylation and impair replication in SARS-CoV-2 infected lung epithelial cells in a cell-type–dependent manner. Targeting GSK-3 may therefore provide an approach to treat COVID-19 and future coronavirus outbreaks.

scholarly journals Targeting the Coronavirus Nucleocapsid Protein through GSK-3 Inhibition

2021 ◽  
Author(s):  
Xiaolei Liu ◽  
Anurag Verma ◽  
Holly Ramage ◽  
Gustavo Garcia ◽  
Rebecca L. Myers ◽  
...  
Keyword(s):  
Nucleocapsid Protein ◽  
Glycogen Synthase ◽  
Lung Epithelial Cells ◽  
Dependent Manner ◽  
Virion Assembly ◽  
N Protein ◽  
Consensus Sequences ◽  
Lung Epithelial ◽  
A Cell ◽  
Transcription And Replication

AbstractThe coronaviruses responsible for severe acute respiratory syndrome (SARS-CoV), COVID-19 (SARS-CoV-2), Middle East respiratory syndrome (MERS-CoV), and other coronavirus infections express a nucleocapsid protein (N) that is essential for viral replication, transcription, and virion assembly. Phosphorylation of N from SARS-CoV by glycogen synthase kinase 3 (GSK-3) is required for its function and inhibition of GSK-3 with lithium impairs N phosphorylation, viral transcription, and replication. Here we report that the SARS-CoV-2 N protein contains GSK-3 consensus sequences and that this motif is conserved in diverse coronaviruses, despite limited overall sequence conservation, raising the possibility that SARS- CoV-2 may be sensitive to GSK-3 inhibitors including lithium. We conducted a retrospective analysis of lithium use in patients from three major health systems who were PCR tested for SARS-CoV-2. We found that patients taking lithium have a significantly reduced risk of COVID- 19 (odds ratio = 0.51 [0.34 - 0.76], p = 0.001). We also show that the SARS-CoV-2 N protein is phosphorylated by GSK-3. Knockout of GSK3A and GSK3B demonstrates that GSK-3 is essential for N phosphorylation. Alternative GSK-3 inhibitors block N phosphorylation and impair replication in SARS-CoV-2 infected lung epithelial cells in a cell-type dependent manner.Targeting GSK-3 may therefore provide a new approach to treat COVID-19 and future coronavirus outbreaks.

scholarly journals Cytotoxic Effects of Air Freshener Biocides in Lung Epithelial Cells

2013 ◽  
Vol 8 (9) ◽  
pp. 1934578X1300800
Author(s):  
Jung-Taek Kwon ◽  
Mimi Lee ◽  
Gun-Baek Seo ◽  
Hyun-Mi Kim ◽  
Ilseob Shim ◽  
...  
Keyword(s):  
Dna Damage ◽  
Epithelial Cells ◽  
Cell Viability ◽  
Lung Epithelial Cells ◽  
Ros Generation ◽  
Dependent Manner ◽  
Cytotoxic Effects ◽  
Lung Epithelial ◽  
Dose Dependent ◽  
A549 Lung

This study evaluated the cytotoxicity of mixtures of citral (CTR) and either benzisothiazolinone (BIT, Mix-CTR-BIT) or triclosan (TCS, Mix-CTR-TCS) in human A549 lung epithelial cells. We investigated the effects of various mix ratios of these common air freshener ingredients on cell viability, cell proliferation, reactive oxygen species (ROS) generation, and DNA damage. Mix-CTR-BIT and Mix-CTR-TCS significantly decreased the viability of lung epithelial cells and inhibited cell growth in a dose-dependent manner. In addition, both mixtures increased ROS generation, compared to that observed in control cells. In particular, cell viability, growth, and morphology were affected upon increase in the proportion of BIT or TCS in the mixture. However, comet analysis showed that treatment of cells with Mix-CTR-BIT or Mix-CTR-TCS did not increase DNA damage. Taken together, these data suggested that increasing the content of biocides in air fresheners might induce cytotoxicity, and that screening these compounds using lung epithelial cells may contribute to hazard assessment.

scholarly journals Inhibition of acute lethal pulmonary inflammation by the IDO–AhR pathway

2017 ◽  
Vol 114 (29) ◽  
pp. E5881-E5890 ◽  
Author(s):  
Soung-Min Lee ◽  
Ha Young Park ◽  
Young-Sill Suh ◽  
Eun Hye Yoon ◽  
Juyang Kim ◽  
...  
Keyword(s):  
T Cells ◽  
Epithelial Cells ◽  
Pulmonary Inflammation ◽  
Lung Parenchyma ◽  
Lung Epithelial Cells ◽  
Dependent Manner ◽  
Independent Manner ◽  
Hematopoietic Stem ◽  
Lung Epithelial ◽  
Ifn Γ

The lung is a prototypic organ that was evolved to reduce immunopathology during the immune response to potentially hazardous endogenous and exogenous antigens. In this study, we show that donor CD4+ T cells transiently induced expression of indoleamine 2,3-dioxygenase (IDO) in lung parenchyma in an IFN-γ–dependent manner early after allogeneic hematopoietic stem cell transplantation (HSCT). Abrogation of host IDO expression by deletion of the IDO gene or the IFN-γ gene in donor T cells or by FK506 treatment resulted in acute lethal pulmonary inflammation known as idiopathic pneumonia syndrome (IPS). Interestingly, IL-6 strongly induced IDO expression in an IFN-γ–independent manner when deacetylation of STAT3 was inhibited. Accordingly, a histone deacetylase inhibitor (HDACi) could reduce IPS in the state where IFN-γ expression was suppressed by FK506. Finally, l-kynurenine produced by lung epithelial cells and alveolar macrophages during IPS progression suppresses the inflammatory activities of lung epithelial cells and CD4+ T cells through the aryl hydrocarbon receptor pathway. Taken together, our results reveal that IDO is a critical regulator of acute pulmonary inflammation and that regulation of IDO expression by HDACi may be a therapeutic approach for IPS after HSCT.

scholarly journals Carbon nanotube uptake changes the biomechanical properties of human lung epithelial cells in a time-dependent manner

2015 ◽  
Vol 3 (19) ◽  
pp. 3983-3992 ◽  
Author(s):  
Chenbo Dong ◽  
Reem Eldawud ◽  
Linda M. Sargent ◽  
Michael L. Kashon ◽  
David Lowry ◽  
...  
Keyword(s):  
Carbon Nanotube ◽  
Exposure Duration ◽  
Human Lung ◽  
Biomechanical Properties ◽  
Engineered Nanomaterials ◽  
Lung Epithelial Cells ◽  
Time Dependent ◽  
Dependent Manner ◽  
Lung Epithelial ◽  
Human Lung Epithelial Cells

The toxicity of engineered nanomaterials in biological systems depends on both the nanomaterial properties and the exposure duration.

scholarly journals SARS-CoV-2 spike protein induces inflammation via TLR2-dependent activation of the NF-κB pathway

2021 ◽  
Author(s):  
Shahanshah Khan ◽  
Mahnoush S. Shafiei ◽  
Christopher Longoria ◽  
John Schoggins ◽  
Rashmin C. Savani ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Inflammatory Response ◽  
Inflammatory Cytokines ◽  
Lung Epithelial Cells ◽  
Dependent Manner ◽  
S Protein ◽  
Cytokines And Chemokines ◽  
Lung Epithelial ◽  
Mouse Macrophages ◽  
Human And Mouse

Pathogenesis of COVID-19 is associated with a hyperinflammatory response; however, the precise mechanism of SARS-CoV-2-induced inflammation is poorly understood. Here we investigated direct inflammatory functions of major structural proteins of SARS-CoV-2. We observed that spike (S) protein potently induces inflammatory cytokines and chemokines including IL-6, IL-1b, TNFa, CXCL1, CXCL2, and CCL2, but not IFNs in human and mouse macrophages. No such inflammatory response was observed in response to membrane (M), envelope (E), and neucleocapsid (N) proteins. When stimulated with extracellular S protein, human lung epithelial cells A549 also produce inflammatory cytokines and chemokines. Interestingly, epithelial cells expressing S protein intracellularly are non-inflammatory, but elicit an inflammatory response in macrophages when co-cultured. Biochemical studies revealed that S protein triggers inflammation via activation of the NF-kB pathway in a MyD88-dependent manner. Further, such an activation of the NF-kB pathway is abrogated in Tlr2-deficient macrophages. Consistently, administration of S protein induces IL-6, TNF-a, and IL-1b in wild-type, but not Tlr2-deficient mice. Together these data reveal a mechanism for the cytokine storm during SARS-CoV-2 infection and suggest that TLR2 could be a potential therapeutic target for COVID-19.

Urokinase induces its own expression in Beas2B lung epithelial cells

2002 ◽  
Vol 283 (2) ◽  
pp. L319-L328 ◽  
Author(s):  
Sreerama Shetty ◽  
Usha R. Pendurthi ◽  
Prathap Kumar Shetty Halady ◽  
Ali O. Azghani ◽  
Steven Idell
Keyword(s):  
Epithelial Cells ◽  
Tyrosine Kinase ◽  
Lung Inflammation ◽  
Cellular Proliferation ◽  
Kinase Inhibitor ◽  
Lung Epithelial Cells ◽  
Cell Surface Receptor ◽  
Dependent Manner ◽  
Amino Terminal ◽  
Lung Epithelial

The urokinase-type plasminogen activator (uPA) interacts with its receptor (uPAR) to promote local proteolysis as well as cellular proliferation and migration. These functions contribute to the pathogenesis of lung inflammation and remodeling as well as the growth and invasiveness of lung neoplasms. In this study, we sought to determine if uPA alters its own expression in lung epithelial cells. Using immunoprecipitation and Western and Northern blotting techniques, we found that uPA treatment enhanced uPA expression in Beas2B lung epithelial cells in a time- and concentration-dependent manner. The induction of uPA expression is mediated through its cell surface receptor uPAR and does not require uPA enzymatic activity. The amino-terminal fragment of uPA, lacking the catalytic domain, is sufficient to induce uPA expression. The serine protease plasmin and the protease inhibitor aprotinin failed to alter uPA-mediated uPA expression, whereas α-thrombin potentiated the response. Pretreatment of Beas2B cells with a tyrosine kinase inhibitor, herbimycin, suggests that activation of tyrosine kinase(s) is involved in the uPA-mediated uPA expression. Induction of uPA expression by exposure of lung-derived epithelial cells to uPA is a newly defined pathway by which this protease could influence expression of local fibrinolytic activity and other uPA-dependent cellular responses germane to lung inflammation or neoplasia.

scholarly journals Cytosolic phospholipase A2α activation induced by S1P is mediated by the S1P3 receptor in lung epithelial cells

2008 ◽  
Vol 295 (2) ◽  
pp. L326-L335 ◽  
Author(s):  
Li-Yuan Chen ◽  
Grzegorz Woszczek ◽  
Sahrudaya Nagineni ◽  
Carolea Logun ◽  
James H. Shelhamer
Keyword(s):  
Epithelial Cells ◽  
Rho Kinase ◽  
A549 Cells ◽  
Lung Epithelial Cells ◽  
Calcium Flux ◽  
Lipid Mediator ◽  
Dependent Manner ◽  
Cytosolic Phospholipase ◽  
Lung Epithelial ◽  
Sphingosine 1 Phosphate

Cytosolic phospholipase A2α (cPLA2α) activation is a regulatory step in the control of arachidonic acid (AA) liberation for eicosanoid formation. Sphingosine 1-phosphate (S1P) is a bioactive lipid mediator involved in the regulation of many important proinflammatory processes and has been found in the airways of asthmatic subjects. We investigated the mechanism of S1P-induced AA release and determined the involvement of cPLA2α in these events in A549 human lung epithelial cells. S1P induced AA release rapidly within 5 min in a dose- and time-dependent manner. S1P-induced AA release was inhibited by the cPLA2α inhibitors methyl arachidonyl fluorophosphonate (MAFP) and pyrrolidine derivative, by small interfering RNA-mediated downregulation of cPLA2α, and by inhibition of S1P-induced calcium flux, suggesting a significant role of cPLA2α in S1P-mediated AA release. Knockdown of the S1P3 receptor, the major S1P receptor expressed on A549 cells, inhibited S1P-induced calcium flux and AA release. The S1P-induced calcium flux and AA release was associated with sphingosine kinase 1 (Sphk1) expression and activity. Furthermore, Rho-associated kinase, downstream of S1P3, was crucial for S1P-induced cPLA2α activation. Our data suggest that S1P acting through S1P3, calcium flux, and Rho kinase activates cPLA2α and releases AA in lung epithelial cells. An understanding of S1P-induced cPLA2α activation mechanisms in epithelial cells may provide potential targets to control inflammatory processes in the lung.

scholarly journals Peptide Mimicrying Between SARS Coronavirus Spike Protein and Human Proteins Reacts with SARS Patient Serum

2008 ◽  
Vol 2008 ◽  
pp. 1-8 ◽  
Author(s):  
K.-Y. Hwa ◽  
W. M. Lin ◽  
Y.-I. Hou ◽  
T.-M. Yeh
Keyword(s):  
Sequence Homology ◽  
Molecular Mimicry ◽  
Lung Epithelial Cells ◽  
Host Cells ◽  
General Strategy ◽  
Sars Coronavirus ◽  
S Protein ◽  
Lung Epithelial ◽  
Human Proteins ◽  
A Cell

Molecular mimicry, defined as similar structures shared by molecules from dissimilar genes or proteins, is a general strategy used by pathogens to infect host cells. Severe acute respiratory syndrome (SARS) is a new human respiratory infectious disease caused by SARS coronavirus (SARS-CoV). The spike (S) protein of SARS-CoV plays an important role in the virus entry into a cell. In this study, eleven synthetic peptides from the S protein were selected based on its sequence homology with human proteins. Two of the peptides D07 (residues 927–937) and D08 (residues 942–951) were recognized by the sera of SARS patients. Murine hyperimmune sera against these peptides bound to proteins of human lung epithelial cells A549. Another peptide D10 (residues 490–502) stimulated A549 to proliferate and secrete IL-8. The present results suggest that the selected S protein regions, which share sequence homology with human proteins, may play important roles in SARS-CoV infection.

scholarly journals Homotypic Interaction of Bunyamwera Virus Nucleocapsid Protein

2005 ◽  
Vol 79 (20) ◽  
pp. 13166-13172 ◽  
Author(s):  
Vincent H. J. Leonard ◽  
Alain Kohl ◽  
Jane C. Osborne ◽  
Angela McLees ◽  
Richard M. Elliott
Keyword(s):  
Nucleocapsid Protein ◽  
Cross Linking ◽  
Genomic Rna ◽  
Rna Dependent Rna Polymerase ◽  
N Protein ◽  
Two Hybrid ◽  
Chemical Cross Linking ◽  
Bunyamwera Virus ◽  
Transcription And Replication ◽  
Weight Structures

ABSTRACT The bunyavirus nucleocapsid protein, N, plays a central role in viral replication in encapsidating the three genomic RNA segments to form functional templates for transcription and replication by the viral RNA-dependent RNA polymerase. Here we report functional mapping of interacting domains of the Bunyamwera orthobunyavirus N protein by yeast and mammalian two-hybrid systems, immunoprecipitation experiments, and chemical cross-linking studies. N forms a range of multimers from dimers to high-molecular-weight structures, independently of the presence of RNA. Deletion of the N- or C-terminal domains resulted in loss of activity in a minireplicon assay and a decreased capacity for N to form higher multimers. Our data suggest a head-to-head and tail-to-tail multimerization model for the orthobunyavirus N protein.

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